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Amaç: Tanı testleri, hasta ve sağlıklı bireylerin oluşturduğu heterojen bir kitlede bireyin gerçek durumunu ortaya çıkarmak amacıyla kullanılır. Bu çalışmada, altın standart test yokluğunda, Bayesci yaklaşım ile ilgilenilen tanı testine ait doğruluk ölçütlerini elde etmek ve diğer yöntemlerle karşılaştırması amaçlanmıştır. Gereç ve Yöntem: Helicobacter Pylori tanısında; HpSA, Kültür, CLO, Histoloji ve PCR gibi farklı testler kullanılmaktadır. Bu çalışmada, Bayesci yaklaşımla parametre tahminlerinin elde edilmesinde önsel bilgileri (prevalans, duyarlılık vs.) içeren modeller oluşturulmuştur. Çalışmada bu tanı testlerinin doğruluklarının saptanmasında ortaya çıkabilen kesin olmayan altın standart yanlılığının düzeltilmesi amacıyla Bayesci yöntemler kullanılmıştır. Bu amaçla H. Pylori ile ilgili bir uzmanlık tezine ait veriler WinBUGS ve R paket programları yardımıyla analiz edilmiştir. Bulgular: Bayesci çıkarsama yapıldığında hastalık ile ilgili prevalans bilgisi ve önsel bilgi dikkate alındığından tanı testine ait pozitif ve negatif kestirim değerlerine ait sonuçların daha güvenilir olduğu saptanmıştır. Kestirim değerlerine benzer şekilde duyarlılık ve seçicilik değerleri içinde güvenilir sonuçlar elde edilmiştir. Bayesci yaklaşım ile elde edilen güvenilir aralık önsel bilgi kullanıldığında daralmıştır. Sonuç: Altın standardın olmadığı durumlarda prevalans gibi önsel bilgileri kullanan Bayesci yaklaşımlar klinisyenler için tanıda kolaylık sağlayacaktır.

Purpose: Diagnostic tests are used for defining the disease status of person in a heterogeneous population which consists of healthy and diseased people. In this study, it was aimed to obtain the accuracy measures of the diagnostic test with the Bayesian approach when there was no gold standard test, and to compare it with other methods. Materials and Methods: In the diagnosis of Helicobacter Pylori, different tests are used such as HpSA, Culture, CLO, Histology and PCR. In this study, models containing a priori information (prevalence, sensitivity, etc.) were created to obtain parameter estimations with the Bayesian approach. Bayesian methods were used to correct the imperfect gold standard bias that may occur in determining the accuracy of these diagnostic tests. For this purpose, the data of a medical thesis on H. Pylori were analyzed with the help of WinBUGS and R package programs. Results: When Bayesian inference is made, the results of the positive and negative predictive values of the diagnostic test were found to be more reliable, since the prevalence and a priori information about the disease were taken into account. Similar to the predictive values, reliable results were obtained for sensitivity and specificity values. The credible interval obtained by using Bayesian approach is narrowed when a prior information is used. Conclusion: In the absence of a gold standard, Bayesian approaches using a prior information such as prevalence and diagnostic test information could facilitate diagnosis for clinicians. [ABSTRACT FROM AUTHOR]

BackgroundBardet-Biedl syndrome (BBS) is a recessive and genetically heterogeneous ciliopathy characterised by retinitis pigmentosa, obesity, kidney dysfunction, postaxial polydactyly, behavioural dysfunction and hypogonadism. 7 of the 17 BBS gene products identified to date assemble together with the protein BBIP1/BBIP10 into the BBSome, a protein complex that ferries signalling receptors to and from cilia.Methods and resultsExome sequencing performed on a sporadic BBS case revealed for the first time a homozygous stop mutation (NM_001195306: c.173T>G, p.Leu58*) in the BBIP1 gene. This mutation is pathogenic since no BBIP1 protein could be detected in fibroblasts from the patient, and BBIP1[Leu58*] is unable to associate with the BBSome subunit BBS4.ConclusionsThese findings identify BBIP1 as the 18th BBS gene (BBS18) and suggest that BBSome assembly may represent a unifying pathomechanism for BBS.

The temperature of the exhaust gas of a diesel piston engine, measured in the characteristic control sections of its thermo-flow system, can be a valuable source of diagnostic information about the technical condition of the elements limiting the working spaces thus separated, including the turbocharging system, but also its fuel supply system and replacement of the medium. In standard marine engine measurement systems equipped with an impulse turbocharging system, the exhaust gas temperature is measured at the outlet of individual cylinders and before and after the turbocharger turbine, using traditional thermocouples with high measurement inertia (time constant of tenths of a second and more). This means that for further diagnostic analyses, the average value of the periodically changing temperature of the exhaust stream leaving individual engine cylinders, the exhaust stream in the collective duct feeding the turbine and the exhaust stream in the exhaust duct of the turbine is used. This article proposes a new approach to the issue of diagnostic informationiveness of the exhaust gas temperature of a diesel engine, extending its observations with the dynamics of changes in the duration of one working cycle. The aim of the tests carried out on the laboratory stand of Farymann Diesel engine type D10 was to determine the diagnostic relations between the loss of permeability of the inlet air channel filter baffle and selected standards of the quick-changing signal of the exhaust gas temperature. On the basis of the calculations carried out, the following dynamic features of the recorded signal were determined: maximum amplitude of instantaneous exhaust gas temperature values (peak-to-peak value), its rate of increase and decrease, and the specific enthalpy of exhaust gases within one engine work cycle. Comparative analysis of numerical data characterizing the recorded quick-changing exhaust gas temperature courses clearly indicates obvious thermodynamic and energy consequences of partial loss of flow capacity of the air channel supplying the combustion chamber of the test engine. A further development of the experimental test programme is foreseen in order to determine a diagnostic matrix to support the diagnostic inference about the technical condition of the diesel engine on the basis of measurements and analysis of the quick-changing exhaust gas temperature.

ABSTRACT Missense, iso-semantic, and intronic mutations are challenging for interpretation, in particular for their impact in m RNA. Various tools such as the Human Splicing Finder ( HSF) system could be used to predict the impact on splicing; however, no diagnosis result could rely on predictions alone, but requires functional testing. Here, we report an in vitro approach to study the impact of DYSF mutations on splicing. It was evaluated on a series of 45 DYSF mutations, both intronic and exonic. We confirmed splicing alterations for all intronic mutations localized in 5′ or 3′ splice sites. Then, we showed that DYSF missense mutations could also result in splicing defects: mutations c.463 G> A and c.2641 A> C abolished ESEs and led to exon skipping; mutations c.565 C> G and c.1555 G> A disrupted Exonic Splicing Enhancer (ESE), while concomitantly creating new 5′ or 3′ splice site leading to exonic out of frame deletions. We demonstrated that 20% of DYSF missense mutations have a strong impact on splicing. This minigene strategy is an efficient tool for the detection of splicing defects in dysferlinopathies, which could allow for a better comprehension of splicing defects due to mutations and could improve prediction tools evaluating splicing defects. [ABSTRACT FROM AUTHOR]

Background Patients with cystic fibrosis (CF) manifest a multisystem disease due to deleterious mutations in each gene encoding the cystic fibrosis transmembrane conductance regulator (). However, the role of dysfunctional CFTR is uncertain in individuals with mild forms of CF (ie, pancreatic sufficiency) and mutation in only one gene. Methods Eleven pancreatic sufficient (PS) CF patients with only one mutation identified after mutation screening (three patients), mutation scanning (four patients) or DNA sequencing (four patients) were studied. Bi-directional sequencing of the coding region of was performed in patients who had mutation screening or scanning. If a second mutation was not identified, mRNA transcripts from nasal epithelial cells were analysed to determine if any PS-CF patients harboured a second mutation that altered RNA expression. Results Sequencing of the coding regions of identified a second deleterious mutation in five of the seven patients who previously had mutation screening or mutation scanning. Five of the remaining six patients with only one deleterious mutation identified in the coding region of one gene had a pathologic reduction in the amount of RNA transcribed from their other gene (8.4–16% of wild type). Conclusions These results show that sequencing of the coding region of followed by analysis of transcription could be a useful diagnostic approach to confirm that patients with mild forms of CF harbour deleterious alterations in both genes. [ABSTRACT FROM PUBLISHER]

Background Fanconi anaemia (FA) is a rare syndrome characterized by bone marrow failure, malformations and cancer predisposition. Chromosome fragility induced by DNA interstrand crosslink (ICL)-inducing agents such as diepoxybutane (DEB) or mitomycin C (MMC) is the ‘gold standard’ test for the diagnosis of FA. Objective To study the variability, the diagnostic implications and the clinical impact of chromosome fragility in FA. Methods Data are presented from 198 DEB-induced chromosome fragility tests in patients with and without FA where information on genetic subtype, cell sensitivity to MMC and clinical data were available. Results This large series allowed quantification of the variability and the level of overlap in ICL sensitivity among patients with FA and the normal population. A new chromosome fragility index is proposed that provides a cut-off diagnostic level to unambiguously distinguish patients with FA, including mosaics, from non-FA individuals. Spontaneous chromosome fragility and its correlation with DEB-induced fragility was also analysed, indicating that although both variables are correlated, 54% of patients with FA do not have spontaneous fragility. The data reveal a correlation between malformations and sensitivity to ICL-inducing agents. This correlation was also statistically significant when the analysis was restricted to patients from the FA-A complementation group. Finally, chromosome fragility does not correlate with the age of onset of haematological disease. Conclusions This study proposes a new chromosome fragility index and suggests that genome instability during embryo development may be related to malformations in FA, while DEB-induced chromosome breaks in T cells have no prognostic value for the haematological disease. [ABSTRACT FROM PUBLISHER]

Background Leber congenital amaurosis (LCA) is the earliest and most severe inherited retinal degeneration. Isolated forms of LCA frequently result from mutation of the gene which is expressed in various ciliated tissues. Methods Seven LCA patients with mutations were investigated to study otorhinolaryngologic phenotype and respiratory cilia. Nasal biopsies and brushing were performed to study cilia ultrastructure using transmission electron microscopy and ciliary beating using high-speed videomicroscopy, respectively. expression in normal nasal epithelium was studied using real-time RT-PCR. Results When electron microscopy was feasible (5/7), high levels of respiratory cilia defects were detected. The main defects concerned dynein arms, central complex and/or peripheral microtubules. All patients had a rarefaction of ciliated cells and a variable proportion of short cilia. Frequent but moderate and heterogeneous clinical and ciliary beating abnormalities were found. was highly expressed in the neural retina and nasal epithelial cells compared with other tissues. Discussion These data provide the first clear demonstration of respiratory cilia ultrastructural defects in LCA patients with mutations. The frequency of these findings in LCA patients along with the high expression of in nasal epithelium suggest that CEP290 has an important role in the proper development of both the respiratory ciliary structures and the connecting cilia of photoreceptors. The presence of respiratory symptoms in patients could represent additional clinical criteria to direct genotyping of patients affected with the genetically heterogeneous cone-rod dystrophy subtype of LCA. [ABSTRACT FROM PUBLISHER]

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Background: Exome and genome sequencing have been demonstrated to increase diagnostic yield in paediatric populations, improving treatment options and providing risk information for relatives. There are limited studies examining the clinical utility of these tests in adults, who currently have limited access to this technology., Methods: Patients from adult and cancer genetics clinics across Toronto, Ontario, Canada were recruited into a prospective cohort study evaluating the diagnostic utility of exome and genome sequencing in adults. Eligible patients were ≥18 years of age and suspected of having a hereditary disorder but had received previous uninformative genetic test results. In total, we examined the diagnostic utility of exome and genome sequencing in 47 probands and 34 of their relatives who consented to participate and underwent exome or genome sequencing., Results: Overall, 17% (8/47) of probands had a pathogenic or likely pathogenic variant identified in a gene associated with their primary indication for testing. The diagnostic yield for patients with a cancer history was similar to the yield for patients with a non-cancer history (4/18 (22%) vs 4/29 (14%)). An additional 24 probands (51%) had an inconclusive result. Secondary findings were identified in 10 patients (21%); three had medically actionable results., Conclusions: This study lends evidence to the diagnostic utility of exome or genome sequencing in an undiagnosed adult population. The significant increase in diagnostic yield warrants the use of this technology. The identification and communication of secondary findings may provide added value when using this testing modality as a first-line test., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. No commercial re-use. See rights and permissions. Published by BMJ.)

Competing Interests: Competing interests: None declared.

PurposeThe purpose of this document is to provide guidance for the use of next-generation sequencing (NGS, also known as massively parallel sequencing or MPS) in Canadian clinical genetic laboratories for detection of genetic variants in genomic DNA and mitochondrial DNA for inherited disorders, as well as somatic variants in tumour DNA for acquired cancers. They are intended for Canadian clinical laboratories engaged in developing, validating and using NGS methods. METHODS OF STATEMENT DEVELOPMENT: The document was drafted by the Canadian College of Medical Geneticists (CCMG) Ad Hoc Working Group on NGS Guidelines to make recommendations relevant to NGS. The statement was circulated for comment to the CCMG Laboratory Practice and Clinical Practice committees, and to the CCMG membership. Following incorporation of feedback, the document was approved by the CCMG Board of Directors. DISCLAIMER: The CCMG is a Canadian organisation responsible for certifying medical geneticists and clinical laboratory geneticists, and for establishing professional and ethical standards for clinical genetics services in Canada. The current CCMG Practice Guidelines were developed as a resource for clinical laboratories in Canada and should not be considered to be inclusive of all information laboratories should consider in the validation and use of NGS for a clinical laboratory service., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Progression time from islet autoimmunity to clinical type 1 diabetes is highly variable and the extent that genetic factors contribute is unknown., Methods: In 341 islet autoantibody-positive children with the human leucocyte antigen (HLA) DR3/DR4-DQ8 or the HLA DR4-DQ8/DR4-DQ8 genotype from the prospective TEDDY (The Environmental Determinants of Diabetes in the Young) study, we investigated whether a genetic risk score that had previously been shown to predict islet autoimmunity is also associated with disease progression., Results: Islet autoantibody-positive children with a genetic risk score in the lowest quartile had a slower progression from single to multiple autoantibodies (p=0.018), from single autoantibodies to diabetes (p=0.004), and by trend from multiple islet autoantibodies to diabetes (p=0.06). In a Cox proportional hazards analysis, faster progression was associated with an increased genetic risk score independently of HLA genotype (HR for progression from multiple autoantibodies to type 1 diabetes, 1.27, 95% CI 1.02 to 1.58 per unit increase), an earlier age of islet autoantibody development (HR, 0.68, 95% CI 0.58 to 0.81 per year increase in age) and female sex (HR, 1.94, 95% CI 1.28 to 2.93)., Conclusions: Genetic risk scores may be used to identify islet autoantibody-positive children with high-risk HLA genotypes who have a slow rate of progression to subsequent stages of autoimmunity and type 1 diabetes., Competing Interests: Competing interests: A patent has been applied for (LU100334) with the title ‘Method the risk to develop type 1 diabetes’ by Helmholtz Zentrum München Deutsches Forschungszentrum für Gesundheit und Umwelt. EB, A-GZ and CW are among the inventors. The patent includes the genetic score that is examined in the manuscript., (© Author(s) (or their employer(s)) 2019. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Advances in molecular technologies and in-silico variant prediction tools offer wide-ranging opportunities in diagnostic settings, yet they also present with significant limitations., Objective: Here, we contextualise the limitations of next-generation sequencing (NGS), multiplex ligation-dependent probe amplification (MLPA) and in-silico prediction tools routinely used by diagnostic laboratories by reviewing specific experiences from our diagnostic laboratory., Methods: We investigated discordant annotations and/or incorrect variant 'callings' in exons of 56 genes constituting our cardiomyopathy and connective tissue disorder NGS panels. Discordant variants and segmental duplications (SD) were queried using the National Center for Biotechnology Information (NCBI) Basic Local Alignment Search Tool and the University of California Santa Cruz genome browser, respectively, to identify regions of high homology. Discrepant variant analyses by in-silico models were re-evaluated using updated file entries., Results: We observed a 5% error rate in MYH7 variant 'calling' using MLPA, which resulted from >90% homology of the MYH7 probe-binding site to MYH6 . SDs were detected in TTN , PKP2 and MYLK . SDs in MYLK presented the highest risk (15.7%) of incorrect variant 'calling'. The inaccurate 'callings' and discrepant in-silico predictions were resolved following detailed investigation into the source of error., Conclusion: Recognising the limitations described here may help avoid incorrect diagnoses and leverage the power of new molecular technologies in diagnostic settings., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Obesity is a global and severe health problem. Due to genetic heterogeneity, the identification of genetic defects in patients with obesity can be time consuming and costly. Therefore, we developed a custom diagnostic targeted next-generation sequencing (NGS)-based analysis to simultaneously identify mutations in 52 obesity-related genes. The aim of this study was to assess the diagnostic yield of this approach in patients with suspected genetic obesity., Methods: DNA of 1230 patients with obesity (median BMI adults 43.6 kg/m 2 ; median body mass index-SD children +3.4 SD) was analysed in the genome diagnostics section of the Department of Genetics of the UMC Utrecht (The Netherlands) by targeted analysis of 52 obesity-related genes., Results: In 48 patients pathogenic mutations confirming the clinical diagnosis were detected. The majority of these were observed in the MC4R gene (18/48). In an additional 67 patients a probable pathogenic mutation was identified, necessitating further analysis to confirm the clinical relevance., Conclusions: NGS-based gene panel analysis in patients with obesity led to a definitive diagnosis of a genetic obesity disorder in 3.9% of obese probands, and a possible diagnosis in an additional 5.4% of obese probands. The highest yield was achieved in a selected paediatric subgroup, establishing a definitive diagnosis in 12 out of 164 children with severe early onset obesity (7.3%). These findings give a realistic insight in the diagnostic yield of genetic testing for patients with obesity and could help these patients to receive (future) personalised treatment., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2018. No commercial re-use. See rights and permissions. Published by BMJ.)

Fetal hypokinesia or akinesia encompasses a broad spectrum of disorders, united by impaired movement in utero. Often, the underlying aetiology is genetic in origin, affecting part of the neuromuscular system. The affordable and high-throughput nature of next-generation DNA sequencing has led to an explosion in disease gene discovery across rare diseases, including fetal akinesias. A genetic diagnosis has clinical utility as it may affect management and prognosis and informs recurrence risk, facilitating family planning decisions. More broadly, knowledge of disease genes increasingly allows population-based preconception carrier screening, which has reduced the incidence of recessive diseases in several populations. Despite gains in knowledge of the genetics of fetal akinesia, many families lack a genetic diagnosis. In this review, we describe the developments in Mendelian genetics of neuromuscular fetal akinesia in the genomics era. We examine genetic diagnoses with neuromuscular causes, specifically including the lower motor neuron, peripheral nerve, neuromuscular junction and muscle., Competing Interests: Competing interests: None declared., (© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)

Genetic testing for hereditary cancer predisposition has evolved rapidly in recent years with the discovery of new genes, but there is much debate over the clinical utility of testing genes for which there are currently limited data regarding the degree of associated cancer risk. To address the discrepancies that have arisen in the provision of these tests across the UK, the UK Cancer Genetics Group facilitated a 1-day workshop with representation from the majority of National Health Service (NHS) clinical genetics services. Using a preworkshop survey followed by focused discussion of genes without prior majority agreement for inclusion, we achieved consensus for panels of cancer genes with sufficient evidence for clinical utility, to be adopted by all NHS genetics services. To support consistency in the delivery of these tests and advice given to families across the country, we also developed management proposals for individuals who are found to have pathogenic mutations in these genes. However, we fully acknowledge that the decision regarding what test is most appropriate for an individual family rests with the clinician, and will depend on factors including specific phenotypic features and the family structure., Competing Interests: Competing interests: None declared., (© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)

Background: Facioscapulohumeral muscular dystrophy (FSHD) is associated with an epigenetic defect on 4qter. Two clinically indistinguishable forms of FSHD are known, FSHD1 and FSHD2. FSHD1 is caused by contraction of the highly polymorphic D4Z4 macrosatellite repeat array on chromosome 4q35. FSHD2 is caused by pathogenic mutations of the SMCHD1 gene.Both genetic defects lead to D4Z4 DNA hypomethylation. In the presence of a polymorphic polyadenylation signal (PAS), DNA hypomethylation leads to inappropriate expression of the D4Z4-encoded DUX4 transcription factor in skeletal muscle. Currently, hypomethylation is not diagnostic per se because of the interference of non-pathogenic arrays and the lack of information about the presence of DUX4-PAS., Methods: We investigated, by bisulfite sequencing, the DNA methylation levels of the region distal to the D4Z4 array selectively in PAS-positive alleles., Results: Comparison of FSHD1, FSHD2 and Control subjects showed a highly significant difference of methylation levels in all CpGs tested. Importantly, using a cohort of 112 samples, one of these CpGs (CpG6) is able to discriminate the affected individuals with a sensitivity of 0.95 supporting this assay potential for FSHD diagnosis. Moreover, our study showed a relationship between PAS-specific methylation and severity of the disease., Conclusions: These data point to the CpGs distal to the D4Z4 array as a critical region reflecting multiple factors affecting the epigenetics of FSHD. Additionally, methylation analysis of this region allows the establishment of a rapid and sensitive tool for FSHD diagnosis., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/)

Background: SOX11 is a transcription factor proposed to play a role in brain development. The relevance of SOX11 to human developmental disorders was suggested by a recent report of SOX11 mutations in two patients with Coffin-Siris syndrome. Here we further investigate the role of SOX11 variants in neurodevelopmental disorders., Methods: We used array based comparative genomic hybridisation and trio exome sequencing to identify children with intellectual disability who have deletions or de novo point mutations disrupting SOX11. The pathogenicity of the SOX11 mutations was assessed using an in vitro gene expression reporter system. Loss-of-function experiments were performed in xenopus by knockdown of Sox11 expression., Results: We identified seven individuals with chromosome 2p25 deletions involving SOX11. Trio exome sequencing identified three de novo SOX11 variants, two missense (p.K50N; p.P120H) and one nonsense (p.C29*). The biological consequences of the missense mutations were assessed using an in vitro gene expression system. These individuals had microcephaly, developmental delay and shared dysmorphic features compatible with mild Coffin-Siris syndrome. To further investigate the function of SOX11, we knocked down the orthologous gene in xenopus. Morphants had significant reduction in head size compared with controls. This suggests that SOX11 loss of function can be associated with microcephaly., Conclusions: We thus propose that SOX11 deletion or mutation can present with a Coffin-Siris phenotype., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/)

Background: Orodental diseases include several clinically and genetically heterogeneous disorders that can present in isolation or as part of a genetic syndrome. Due to the vast number of genes implicated in these disorders, establishing a molecular diagnosis can be challenging. We aimed to develop a targeted next-generation sequencing (NGS) assay to diagnose mutations and potentially identify novel genes mutated in this group of disorders., Methods: We designed an NGS gene panel that targets 585 known and candidate genes in orodental disease. We screened a cohort of 101 unrelated patients without a molecular diagnosis referred to the Reference Centre for Oro-Dental Manifestations of Rare Diseases, Strasbourg, France, for a variety of orodental disorders including isolated and syndromic amelogenesis imperfecta (AI), isolated and syndromic selective tooth agenesis (STHAG), isolated and syndromic dentinogenesis imperfecta, isolated dentin dysplasia, otodental dysplasia and primary failure of tooth eruption., Results: We discovered 21 novel pathogenic variants and identified the causative mutation in 39 unrelated patients in known genes (overall diagnostic rate: 39%). Among the largest subcohorts of patients with isolated AI (50 unrelated patients) and isolated STHAG (21 unrelated patients), we had a definitive diagnosis in 14 (27%) and 15 cases (71%), respectively. Surprisingly, COL17A1 mutations accounted for the majority of autosomal-dominant AI cases., Conclusions: We have developed a novel targeted NGS assay for the efficient molecular diagnosis of a wide variety of orodental diseases. Furthermore, our panel will contribute to better understanding the contribution of these genes to orodental disease., Trial Registration Numbers: NCT01746121 and NCT02397824., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/)

Background: Countries with culturally accepted consanguinity provide a unique resource for the study of rare recessively inherited genetic diseases. Although hereditary hearing loss (HHL) is not uncommon, it is genetically heterogeneous, with over 85 genes causally implicated in non-syndromic hearing loss (NSHL). This heterogeneity makes many gene-specific types of NSHL exceedingly rare. We sought to define the spectrum of autosomal recessive HHL in Iran by investigating both common and rarely diagnosed deafness-causing genes., Design: Using a custom targeted genomic enrichment (TGE) panel, we simultaneously interrogated all known genetic causes of NSHL in a cohort of 302 GJB2-negative Iranian families., Results: We established a genetic diagnosis for 67% of probands and their families, with over half of all diagnoses attributable to variants in five genes: SLC26A4, MYO15A, MYO7A, CDH23 and PCDH15. As a reflection of the power of consanguinity mapping, 26 genes were identified as causative for NSHL in the Iranian population for the first time. In total, 179 deafness-causing variants were identified in 40 genes in 201 probands, including 110 novel single nucleotide or small insertion-deletion variants and three novel CNV. Several variants represent founder mutations., Conclusion: This study attests to the power of TGE and massively parallel sequencing as a diagnostic tool for the evaluation of hearing loss in Iran, and expands on our understanding of the genetics of HHL in this country. Families negative for variants in the genes represented on this panel represent an excellent cohort for novel gene discovery., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/)

Background: Phenotypic overlap among the inherited bone marrow failure syndromes (IBMFSs) frequently limits the ability to establish a diagnosis based solely on clinical features. >70 IBMFS genes have been identified, which often renders genetic testing prolonged and costly. Since correct diagnosis, treatment and cancer surveillance often depend on identifying the mutated gene, strategies that enable timely genotyping are essential., Methods: To overcome these challenges, we developed a next-generation sequencing assay to analyse a panel of 72 known IBMFS genes. Cases fulfilling the clinical diagnostic criteria of an IBMFS but without identified causal genotypes were included., Results: The assay was validated by detecting 52 variants previously found by Sanger sequencing. A total of 158 patients with unknown mutations were studied. Of 75 patients with known IBMFS categories (eg, Fanconi anaemia), 59% had causal mutations. Among 83 patients with unclassified IBMFSs, we found causal mutations and established the diagnosis in 18% of the patients. The assay detected mutant genes that had not previously been reported to be associated with the patient phenotypes. In other cases, the assay led to amendments of diagnoses. In 20% of genotype cases, the results indicated a cancer surveillance programme., Conclusions: The novel assay is efficient, accurate and has a major impact on patient care., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Background: The Canadian Open Genetics Repository is a collaborative effort for the collection, storage, sharing and robust analysis of variants reported by medical diagnostics laboratories across Canada. As clinical laboratories adopt modern genomics technologies, the need for this type of collaborative framework is increasingly important., Methods: A survey to assess existing protocols for variant classification and reporting was delivered to clinical genetics laboratories across Canada. Based on feedback from this survey, a variant assessment tool was made available to all laboratories. Each participating laboratory was provided with an instance of GeneInsight, a software featuring versioning and approval processes for variant assessments and interpretations and allowing for variant data to be shared between instances. Guidelines were established for sharing data among clinical laboratories and in the final outreach phase, data will be made readily available to patient advocacy groups for general use., Results: The survey demonstrated the need for improved standardisation and data sharing across the country. A variant assessment template was made available to the community to aid with standardisation. Instances of the GeneInsight tool were provided to clinical diagnostic laboratories across Canada for the purpose of uploading, transferring, accessing and sharing variant data., Conclusions: As an ongoing endeavour and a permanent resource, the Canadian Open Genetics Repository aims to serve as a focal point for the collaboration of Canadian laboratories with other countries in the development of tools that take full advantage of laboratory data in diagnosing, managing and treating genetic diseases., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Background: Alport syndrome is a clinically heterogeneous, progressive nephropathy caused by mutations in collagen IV genes, namely COL4A3 and COL4A4 on chromosome 2 and COL4A5 on chromosome X. The wide phenotypic variability and the presence of incomplete penetrance suggest that a simple Mendelian model cannot completely explain the genetic control of this disease. Therefore, we explored the possibility that Alport syndrome is under digenic control., Methods: Using massively parallel sequencing, we identified 11 patients who had pathogenic mutations in two collagen IV genes. For each proband, we ascertained the presence of the same mutations in up to 12 members of the extended family for a total of 56 persons studied., Results: Overall, 23 mutations were found. Individuals with two pathogenic mutations in different genes had a mean age of renal function deterioration intermediate with respect to the autosomal-dominant form and the autosomal-recessive one, in line with molecule stoichiometry of the disruption of the type IV collagen triple helix., Conclusions: Segregation analysis indicated three possible digenic segregation models: (i) autosomal inheritance with mutations on different chromosomes, resembling recessive inheritance (five families); (ii) autosomal inheritance with mutations on the same chromosome resembling dominant inheritance (two families) and (iii) unlinked autosomal and X-linked inheritance having a peculiar segregation (four families). This pedigree analysis provides evidence for digenic inheritance of Alport syndrome. Clinical geneticists and nephrologists should be aware of this possibility in order to more accurately assess inheritance probabilities, predict prognosis and identify other family members at risk., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Background Previous publications and utilisation of risk models for BRCA1 and BRCA2 mutation identification suggests that multiple primary disease in an individual is a strong predictor of a BRCA1/2 mutation and that this is more predictive than the same cancers occurring in close relatives. Methods This study assessed the pathological mutation detection rates for BRCA1, BRCA2 and the CHEK2c.1100 delC mutation in 2022 women with breast cancer, including 100 with breast/ovary double primary and 255 with bilateral breast cancer. Results and discussion Although detection rates for mutations in BRCA1/2 are high at 49% for breast/ovarian double primary and 34% for bilateral breast cancer, the differential effect of multiple primaries in an individual appears to have been overestimated, particularly in those families with only a few malignancies. Nonetheless, bilateral breast cancer does differentially enhance detection rates in strong familial aggregations. CHEK2 1100 DelC mutation rates were lower in bilateral than for unilateral cases at 0.8% compared to 2%. The detected mutation rates for isolated double primary breast and ovarian cancer was 14% (3/22) compared to 17% (17/99) for the same two primaries in two close relatives in families with no other cases of breast/ovarian cancer. Risk models may need to be adjusted if further studies corroborate these findings. [ABSTRACT FROM AUTHOR]

Background Cohen syndrome is a rare autosomal recessive inherited disorder that results from mutations of the VPS13B gene. Clinical features consist of a combination of mental retardation, facial dysmorphism, postnatal microcephaly, truncal obesity, slender extremities, joint hyperextensibility, myopia, progressive chorioretinal dystrophy, and intermittent neutropenia. Patients and methods The aim of the study was to determine which of the above clinical features were the best indicators for the presence of VPS13B gene mutations in a series of 34 patients with suspected Cohen syndrome referred for molecular analysis of VPS13B. Results 14 VPS13B gene mutations were identified in 12 patients, and no mutation was found in 22 patients. The presence of chorioretinal dystrophy (92% vs 32%, p=0.0023), intermittent neutropenia (92% vs 5%, p<0.001), and postnatal microcephaly (100% vs 48%, p=0.0045) was significantly higher in the group of patients with a VPS13B gene mutation compared to the group of patients without a mutation. All patients with VPS13B mutations had chorioretinal dystrophy and/or intermittent neutropenia. The Kolehmainen diagnostic criteria provided 100% sensibility and 77% specificity when applied to this series. Conclusion From this study and a review of more than 160 genotyped cases from the literature, it is concluded that, given the large size of the gene, VPS13B screening is not indicated in the absence of chorioretinal dystrophy or neutropenia in patients aged over 5 years. The follow-up of young patients could be a satisfactory alternative unless there are some reproductive issues. [ABSTRACT FROM AUTHOR]

Background Birt-Hogg-Dubèc) syndrome (BHDS) is an inherited autosomal genodermatosis characterised by fibrofolliculomas of the skin, renal tumours and multiple lung cysts. Genetic studies have disclosed that the clinical picture as well as responsible germline FLCN mutations are diverse. Objectives BHDS may be caused by a germline deletion which cannot be detected by a conventional genetic approach. Real-time quantitative polymerase chain reaction (qPCR) may be able to identify such a mutation and thus provide us with a more accurate clinical picture of BHDS. Methods This study analysed 36 patients with multiple lung cysts of undetermined causes. Denaturing high performance liquid chromatography (DHPLC) was applied for mutation screening. If no abnormality was detected by DHPLC, the amount of each FLCN exon in genome was quantified by qPCR. Results An FLCN germline mutation was found in 23 (63.9%) of the 36 patients by DHPLC and direct sequencing (13 unique small nucleotide alterations which included 11 novel mutations). A large genomic deletion was identified in two of the remaining 13 patients by qPCR (one patient with exon 14 deletion and one patient with a deletion encompassing exons 9 to 14). Mutations including genomic deletions were most frequently identified in the 3'-end of the FLCN gene including exons 12 and 13 (13/25=52.0%). The BHDS patients whose multiple cysts prompted the diagnosis in this study showed a very low incidence of skin and renal involvement. Conclusions BHDS is due to large deletions as well as small nucleotide alterations. Racial differences may occur between Japanese and patients of European decent in terms of FLCN mutations and clinical manifestations. [ABSTRACT FROM AUTHOR]